The potential for utilizing biological systems as a source of biodegradable thermoplastics is becoming increasingly attractive given the problems with disposal of traditional oil based polymers. Biodegradable polymers are now being investigated as potential vehicles for controlled release of drugs, artificial skin and sutures. Polyhydroxyalkanoates (PHAs) are natural products which can constitute up to 80% of the dry cell weight of bacteria, when the bacteria are subjected to deprivation of O2, nitrogen, phosphates. These biological polyesters are composed of a variety of 3R hydroxyfatty acids (from C4 to C12) resulting in polymers of a wide range of molecular weights, depending on the organism and the C- source available for growth. We have recently cloned, sequenced, overexpressed, and purified the PHA synthase from Alcaligenes eutrophus. This enzyme catalyzes formation in vivo of polyhydroxybutyrates (PHBs) and polyhydroxyvalerates (PHVs). We have also cloned and sequenced the synthase from Pseudomonas oleovorans. Preliminary results suggest that we have been successful in expression of this protein as well. This enzyme polymerizes (C6-C12) 3R-hydroxyfatty acids. The present proposal seeks support for characterization of the A eutrophus synthase as well as purification of the P oleovorans synthase. The substrate specificity of these proteins will be investigated in detail as will their unique mechanisms of initiation, elongation and termination. Both protein analogs (site directed mutants) and substrate analogs (mechanism based inhibitors) will be used as probes. The mechanism of processivity will also be investigated. These studies should provide new insight into the mechanisms of synthases that catalyze formation of homopolymers. The information learned is crucial to the design, using biotechnological advances, of synthases which can make homo and co.polymers which will be useful, due to their biodegradability and unique thermoplastic and elastic properties, to the community in general. Recently these polymers have been found in eucaryotic membranes as well as in human plasma. The function and biosynthesis of these polymers promises to be interesting.